Figure 1.
A set of vectors that drive expression of Cas9 and sgRNAs in C. elegans. (A) The C. elegans eft-3 promoter drives transcription of Cas9 with a 3′ SV40 nuclear localization sequence. A pol III promoter (derived from a U6 snRNA locus) drives transcription of the sgRNA, which contains a target sequence and a scaffold sequence. (B) A schematic illustration of Cas9 interacting with sgRNA and its genomic target. (C) RT-PCR results demonstrating expression of Cas9 and sgRNA transcripts. Total RNA was tested from strains carrying Cas9 vector alone (lanes 1 and 2), unc-119 sgRNA vector alone (lanes 3 and 4), and both vectors (lanes 5 and 6) with primers specific for Cas9 (top panel) or unc-119 sgRNA (bottom panel). For all samples, control reactions were run in the absence of Reverse Transcriptase (-RT; lanes 1, 3, and 5).